AB316924

Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free

Recombinant
RabMAb
KO Validated
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Rabbit Recombinant Monoclonal RIP phospho S166 antibody. Carrier free. Suitable for WB, Dot and reacts with Transfected cell lysate - Human, Human, Synthetic peptide - Human samples.

View Alternative Names

RIP, RIP1, Receptor-interacting serine/threonine-protein kinase 1, Cell death protein RIP, Receptor-interacting protein 1, RIP-1

4 Images

Supplier Data

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

This data was developed using ab316923, the same antibody clone in a different buffer formulation.

The expression of RIP (phospho S166) is upregulated in response to z-VAD, TNF alpha and SM-164, while it is downregulated in response to necrostain treatment (PMID : 32027418, PMID : 33273695).

The identity of the lower MW band at approximately 35 kDa is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-RIP antibody - Total protein control (ab300617) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] (<a href='/en-us/products/primary-antibodies/rip-phospho-s166-antibody-epr25654-166-ab316923'>ab316923</a>) at 1/1000 dilution

Lane 1:

Untreated HT-29 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 40 µg with NFDM/TBST

Lane 2:

HT-29 treated with 20 uM z-VAD for 3.5 hours, then with 20 ng/ml TNF alpha, 100 nM SM-164 and 50 uM necrostatin added for 3 hours whole cell lysate (untreated membrane) at 40 µg with NFDM/TBST

Lane 3:

HT-29 treated with 20 uM z-VAD for 3.5 hours, then with 20 ng/ml TNF alpha and 100nM SM-164 added for 3 hours whole cell lysate (untreated membrane) at 40 µg with NFDM/TBST

Lane 4:

HT-29 treated with 20 uM z-VAD for 3.5 hours, then with 20 ng/ml TNF alpha and 100 nM SM-164 added for 3 hours whole cell lysate (Lambda phosphatase treated membrane) at 40 µg with NFDM/TBST

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 75 kDa,36 kDa

false

Exposure time: 180s

Supplier Data

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

This data was developed using ab316923, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

In Western blot, ab316923 was shown to bind specifically to RIP. Target of interest was observed at 75 kDa in wild-type HAP1 cell lysates (lanes 1 and 3), whereas no signal observed at this size in RIP knockout cell line lysates (lanes 2 and 4).

The expression of RIP (phospho S166) is upregulated in response to z-VAD, TNF alpha and SM-164 treatment (PMID : 32027418, PMID : 33273695).

The identity of the lower MW band at approximately 15 kDa is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-RIP antibody - Total protein control (ab300617) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] (<a href='/en-us/products/primary-antibodies/rip-phospho-s166-antibody-epr25654-166-ab316923'>ab316923</a>) at 1/1000 dilution

Lane 1:

Untreated wild-type HAP1(human chronic myelogenous leukemia near-haploid cell line) whole cell lysate at 50 µg

Lane 2:

Untreated RIP knockout HAP1 whole cell lysate at 50 µg

Lane 3:

Wild-type HAP1 treated with 20 uM z-VAD for 3.5 hours, then with 20 ng/ml TNF alpha and 100 nM SM-164 added for 3 hours whole cell lysate at 50 µg

Lane 4:

RIP knockout HAP1 treated with 20 uM z-VAD for 3.5 hours, then with 20 ng/ml TNF alpha and 100 nM SM-164 added for 3 hours whole cell lysate at 50 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 75 kDa,36 kDa

false

Exposure time: 180s

Supplier Data

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

This data was developed using ab316923, the same antibody clone in a different buffer formulation.

The identity of the lower MW band at approximately 35 kDa is unknown.

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

In Western blot, Anti-RIP antibody - Total protein control (ab300617) staining at 1/1000 dilution.

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] (<a href='/en-us/products/primary-antibodies/rip-phospho-s166-antibody-epr25654-166-ab316923'>ab316923</a>) at 1/1000 dilution

Lane 1:

Untreated 293T (human embryonic kidney epithelial cell) transfected with an empty vector containing a myc-His-tag®, whole cell lysate at 20 µg with NFDM/TBST

Lane 2:

Untreated 293T transfected with a human wild-type RIP expression vector containing a myc-His-tag®, whole cell lysate at 20 µg with NFDM/TBST

Lane 3:

293T transfected with a human wild-type RIP expression vector containing a myc-His-tag® were treated with 100 nM Calyculin for 30 minutes, whole cell lysate at 20 µg with NFDM/TBST

Lane 4:

293T transfected with a human RIP (S166A mutation) expression vector containing a myc-His-tag® were treated with 100 nM Calyculin for 30 minutes, whole cell lysate at 20 µg with NFDM/TBST

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 75 kDa,36 kDa

false

Exposure time: 6s

Supplier Data

Dot Blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

This data was developed using ab316923, the same antibody clone in a different buffer formulation.

Dot blot analysis of RIP (phospho S166) using ab316923 at 1 : 1000 (0.502 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution.

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Dot Blot - Anti-RIP (phospho S166) antibody [EPR25654-166] (<a href='/en-us/products/primary-antibodies/rip-phospho-s166-antibody-epr25654-166-ab316923'>ab316923</a>) at 1/1000 dilution

Lane 1:

RIP (phospho S166) peptide a

Lane 2:

RIP (phospho S166) peptide b

Lane 3:

RIP non-phospho peptide

Secondary

All lanes:

Dot Blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

false

Exposure time: 180s

Unconjugated

Anti-RIP (phospho S166) antibody [EPR25654-166]

Key facts

Host species

Rabbit

Clonality

Monoclonal

Clone number

EPR25654-166

Isotype

IgG

Carrier free

Yes

Reacts with

Human

Applications

Dot, WB

applications

Immunogen

The exact immunogen used to generate this antibody is proprietary information.

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Reactivity data

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Product details

ab316924 is the carrirer-free version of ab316923.

Patented technology
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

What are the advantages of a recombinant monoclonal antibody?
This product is a recombinant monoclonal antibody, which offers several advantages including:

- High batch-to-batch consistency and reproducibility
- Improved sensitivity and specificity
- Long-term security of supply
- Animal-free batch production

For more information, read more on recombinant antibodies.

Conjugation ready
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with 1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

Compatibility
This product is compatible with the Maxpar^® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar^® is a trademark of Fluidigm Canada Inc.

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Properties and storage information

Form

Liquid

Purification technique

Affinity purification Protein A

Storage buffer

pH: 7.2 - 7.4 Constituents: PBS

Shipped at conditions

Blue Ice

Appropriate short-term storage conditions

+4°C

Appropriate long-term storage conditions

+4°C

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Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

RIP also known as Receptor-Interacting Protein or RIPK1 is a serine/threonine-protein kinase with a mass of approximately 74 kDa. It plays an important role in cell death and survival signaling pathways. RIP is expressed ubiquitously across various tissues indicating its importance in many cellular functions. The protein contains a kinase domain an intermediate domain for protein-protein interactions and a death domain which facilitates its involvement in apoptotic signaling processes.

Biological function summary

Receptor-Interacting Protein Kinase 1 (RIPK1) participates in regulating both necroptosis and apoptosis distinguishing itself as an important mediator in cell death mechanisms. As part of the necrosome complex which includes RIPK3 and MLKL RIPK1 functions in necroptosis—a programmed form of necrosis. This characteristic involvement shows its dual role in maintaining cell fate decisions making it an integral part of immune response and inflammation control.

Pathways

RIPK1 strongly associates with the TNF signaling pathway and NF-kB pathway. Its interaction with TNF receptor 1 (TNFR1) and consequent involvement with TRADD and TRAF2 mediates the signal transduction necessary for the activation of NF-kB leading to transcription of genes involved in survival and inflammation. This connection illustrates its capability to switch between promoting cell survival through NF-kB and facilitating cell death via necroptosis or apoptosis depending on cellular context and cues.

RIPK1 plays a significant role in conditions such as inflammatory diseases and neurodegenerative disorders. Its overactivation results in excessive cell death implicated in inflammatory conditions; necrostatin a necroptosis inhibitor targets RIPK1 to potentially mitigate this damage. Furthermore RIPK1's dysregulation links to Alzheimer's disease where it can interact with components like RIPK3 to exacerbate neurodegenerative processes. This relationship underlines the potential of targeting RIPK1 therapeutically to manage inflammation and neurodegeneration.

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Product protocols

For this product, it's our understanding that no specific protocols are required. You can visit:

Visit the General protocols
Visit the Troubleshooting

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Target data

Serine-threonine kinase which is a key regulator of TNF-mediated apoptosis, necroptosis and inflammatory pathways (PubMed : 17703191, PubMed : 24144979, PubMed : 31827280, PubMed : 31827281, PubMed : 32657447, PubMed : 35831301). Exhibits kinase activity-dependent functions that regulate cell death and kinase-independent scaffold functions regulating inflammatory signaling and cell survival (PubMed : 11101870, PubMed : 19524512, PubMed : 19524513, PubMed : 29440439, PubMed : 30988283). Has kinase-independent scaffold functions : upon binding of TNF to TNFR1, RIPK1 is recruited to the TNF-R1 signaling complex (TNF-RSC also known as complex I) where it acts as a scaffold protein promoting cell survival, in part, by activating the canonical NF-kappa-B pathway (By similarity). Kinase activity is essential to regulate necroptosis and apoptosis, two parallel forms of cell death : upon activation of its protein kinase activity, regulates assembly of two death-inducing complexes, namely complex IIa (RIPK1-FADD-CASP8), which drives apoptosis, and the complex IIb (RIPK1-RIPK3-MLKL), which drives necroptosis (By similarity). RIPK1 is required to limit CASP8-dependent TNFR1-induced apoptosis (By similarity). In normal conditions, RIPK1 acts as an inhibitor of RIPK3-dependent necroptosis, a process mediated by RIPK3 component of complex IIb, which catalyzes phosphorylation of MLKL upon induction by ZBP1 (PubMed : 19524512, PubMed : 19524513, PubMed : 29440439, PubMed : 30988283). Inhibits RIPK3-mediated necroptosis via FADD-mediated recruitment of CASP8, which cleaves RIPK1 and limits TNF-induced necroptosis (PubMed : 19524512, PubMed : 19524513, PubMed : 29440439, PubMed : 30988283). Required to inhibit apoptosis and necroptosis during embryonic development : acts by preventing the interaction of TRADD with FADD thereby limiting aberrant activation of CASP8 (By similarity). In addition to apoptosis and necroptosis, also involved in inflammatory response by promoting transcriptional production of pro-inflammatory cytokines, such as interleukin-6 (IL6) (PubMed : 31827280, PubMed : 31827281). Phosphorylates RIPK3 : RIPK1 and RIPK3 undergo reciprocal auto- and trans-phosphorylation (PubMed : 19524513). Phosphorylates DAB2IP at 'Ser-728' in a TNF-alpha-dependent manner, and thereby activates the MAP3K5-JNK apoptotic cascade (PubMed : 15310755, PubMed : 17389591). Required for ZBP1-induced NF-kappa-B activation in response to DNA damage (By similarity).

See full target information RIPK1 phospho S166

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Product promise

We are committed to supporting your work with high-quality reagents, and we're here for you every step of the way. In the unlikely event that one of our products does not perform as expected, you're protected by our Product Promise.

For full details, please see our Terms & Conditions

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

For licensing inquiries, please contact partnerships@abcam.com

Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

Western blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

Dot Blot - Anti-RIP (phospho S166) antibody [EPR25654-166] - BSA and Azide free (AB316924)

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Key facts

Host species

Clonality

Clone number

Isotype

Carrier free

Reacts with

Applications

Immunogen

Reactivity data

Product details

Properties and storage information

Form

Purification technique

Storage buffer

Shipped at conditions

Appropriate short-term storage conditions

Appropriate long-term storage conditions

Supplementary information

Biological function summary

Pathways

Product protocols

Target data

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Primary Antibodies

Secondary Antibodies

Primary Antibodies

Product promise